An understanding of the regulation of protein metabolism in skeletal muscle is required for the development of rational approaches to the treatment of diabetes mellitus and other disease states that affect nutrient homeostasis. Toward this end, the overall goal of this project is to gain a better understanding of the mechanisms by which hormones and nutrients regulate the pathway of protein synthesis in skeletal muscle. The specific focus of the project continues to be on translational control, particularly the regulation of initiation of protein synthesis. In the next grant period, special attention is to be given to the mechanism of action of insulin on specific initiation factors, particularly eIF-2 and eIF-2B. Furthermore, the regulatory role of amino acids is to be investigated because of the recent demonstration of their importance in controlling protein synthesis in skeletal muscle in humans. Additionally, the project is to be extended from skeletal to cardiac muscle so as to better contrast the variations in translational control manifested by muscles composed of different fiber types. The specific aims of the project are: (1) to develop monospecific antibodies and cDNA clones to individual subunits of initiation factors that are found to be important in the regulation of protein synthesis in muscle; (2) to investigate protein and mRNA expression as well as structure/function relationships of specific initation factors in muscle using monospecific antibodies and cDNA clones; (3) to investigate the mechanism by which insulin stimulates initiation of protein synthesis and causes an enhancement of eIF-2B activity in muscle; (4) to identify in skeletal muscle the specific initiation factor(s) involved in regulation of initiation of protein synthesis by amino acids and hormones other than insulin; and (5) to identify in cardiac muscle the specific initiation factor(s) involved in the regulation of initiation of protein synthesis by fatty acids and other oxidizable noncarbohydrate substrates. Overall , the studies described in this proposal should help to identify mechanisms by which hormones and nutrients regulate protein synthesis in skeletal and cardiac muscle. They should also provide new insights into the biochemical and molecular mechanisms involved in the initiation of protein synthesis.